Cytochrome P-450 is the terminal oxidase of one of the monooxygenase systems found in the endoplasmic reticulum of many tissues. This monooxygenase catalyzes the oxidative metabolism of a wide variety of organic compounds including polycyclic aromatic hydrocarbons, fatty acids, steroids and drugs. Most of these reactions, like drug detoxification, are beneficial but some like the epoxidation of polycyclic aromatic hydrocarbons are not because these compounds are then carcinogenic. In no case is the metabolic control of this enzyme system understood. The objective of this research is to study the mechanism and control of cytochrome P-450 catalyzed reactions. The enzyme source used will be the bacterium Pseudomonas putida because its cytochrome P-450 system is soluble and readily purified and has been previously shown to be an excellent model for the comparable mammalian membrane bound enzyme. The work will proceed on four related projects: a) studies of the electron transfer reaction between reduced putidaredoxin and oxidized cytochrome P-450; b) characterization of the electron transfer and oxygen activation reaction; c) purify and characterize the flavoprotein putidaredoxin reductase; and d) develop a computer model which will encompass the reaction dynamics and control of this complex enzyme system. The techniques which will be used in this project include: a) recording spectrophotometry; b) stopped-flow spectrophotometry with kinetic analysis of the data obtained; c) electron paramagnetic resonance spectrometry; and d) computer modeling of metabolic control.